Fig. 6: Inhibition of GLUT1 expression abolishes the metformin-induced increase in glucose homeostasis and glucotonic effect in the distal intestine.

a Post-washing FDG autoradiography of intestinal tissues from vehicle-treated mice, metformin-treated moce and metformin-treated mice receiving intraperitoneal injection of STF-31 (a GLUT1-specific inhibitor). b FDG autoradiography of the relative radiotracer amount in the ileum (IL) and colon (COL) after washing. The STF-31-injected ileum and colon show significantly lower FDG uptake than the metformin-treated ileum and colon. The vehicle-treated ileum and colon were used for comparison. c Colon PBS washing analysis. PBS washings from the colon of STF-31-injected mice showed lower FDG excretion compared with those from metformin-treated mice. The vehicle-treated ileum and colon were used for comparison. d IPGTT data comparing metformin-treated mice, metformin-treated mice with GLUT1 inhibitor (STF-31) intraperitoneal injection and vehicle-treated mice. The metformin group showed significant improvement in glucose tolerance compared with the vehicle group. This improvement in glucose tolerance was significantly reduced in the metformin with STF-31 injection group compared with the metformin group (n = 5, analysis of variance multiple t-test for multiple-comparison correction) and corresponding area under the curves (AUCs). e The expression of GLUT1, HK2 and TXNIP in the ileum and colon from vehicle- and metformin-treated mice. f An immunoblot showing the increased GLUT1 and decreased TXNIP from ileum and colon samples from metformin-treated mice (n = 3 mice per group). Expression levels were normalized to β-actin. g Membrane GLUT1 expression in ileum and colon from vehicle- and metformin-treated mice (n = 3 mice per group). Expression levels were normalized to ATP1A1. h, i Representative images of GLUT1 (h) and TXNIP (i) immunostaining of the ileum and colon from vehicle-treated and metformin-treated C57BL/6 mice. Scale bars, 100 μm (left images) and 50 μm (right images). Statistical comparisons were performed on the percentage of strongly 3,3′-Diaminobenzidine (DAB)-stained area between vehicle- and metformin-treated groups. j, Representative fluorescence images of GLUT1 after vehicle and metformin treatment in human intestinal organoids. Red signals represent phalloidin 4 staining, indicating F-actin fibers. Blue signals represent DAPI staining, indicating nuclei location. Green signals represent GLUT1 expression. Scale bars, 50 μm. All data are presented as the mean ± s.e.m. Data in b–e were analyzed using two-tailed Student’s t-tests; *P < 0.05, **P < 0.01, ***P < 0.001.